EVIDENCE FOR A HIGH-AFFINITY UPTAKE SYSTEM FOR CHOLECYSTOKININ-OCTAPEPTIDE (CCK8) IN RAT CORTICAL SYNAPTOSOMES

Given the high resistance of the cholecystokinin octapeptide (CCK8) to in vivo peptidase degradation, the possible existence of a reuptake s ystem for this peptide was investigated. Efficient accumulation of int act, tritiated propionyl CCK8 ([H-3]pCCK(8)) was observed following it s incubation with rat cortical synaptosomes but not with cerebellar sy naptosomes, where no cholecystokinin immunoreactivity was found. This uptake process appeared to be dependent on temperature, duration of in cubation, concentration of radioligand, the presence of glucose and th e integrity of the synaptosomes. A Lineweaver-Burk analysis indicated that the putative uptake process is characterized by a single K-m valu e of 10.7 nM and a V-max of 8.5 fmol/min/mg of protein. Carbonyl cyani de-m-chlorophenyl hydrazone, an uncoupler of oxidative phosphorylation , blocked accumulation of [H-3]pCCK(8), whereas ouabain did not. The u ptake was found to be highly specific since, among all the cholecystok inin analogues tested, only CCK8 and, to a lesser extent, CCK7, were a ble to inhibit [H-3]pCCK(8) uptake. The rate of [H-3]pCCK(8) uptake wa s not affected by CCK4, CCK5, D-Trp CCK8, BC 264, a potent and selecti ve CCK-B agonist, and L-365,260, a selective CCK-B antagonist. In addi tion, no accumulation of radioactivity was observed using [H-3]pBC 264 , a result which is not in favour of a cholecystokinin receptor-induce d internalization mechanism. The potent and selective uptake mechanism characterized in this study could participate, in conjunction with ex tra and intracellular degradation of CCK8 by peptidases, in the interr uption of cholecystokinin-conveyed messages in the brain.